AB-423 is a member of the sulfamoylbenzamide (SBA) class of hepatitis B virus (HBV) capsid inhibitors in phase 1 clinical trials. In cell culture models, AB-423 showed potent inhibition of HBV replication (50% effective concentration [EC] = 0.08 to 0.27 μM; EC = 0.33 to 1.32 μM) with no significant cytotoxicity (50% cytotoxic concentration > 10 μM). Addition of 40% human serum resulted in a 5-fold increase in the ECs. AB-423 inhibited HBV genotypes A through D and nucleos(t)ide-resistant variants Treatment of HepDES19 cells with AB-423 resulted in capsid particles devoid of encapsidated pregenomic RNA and relaxed circular DNA (rcDNA), indicating that it is a class II capsid inhibitor. In a infection model, AB-423 prevented the conversion of encapsidated rcDNA to covalently closed circular DNA, presumably by interfering with the capsid uncoating process. Molecular docking of AB-423 into crystal structures of heteroaryldihydropyrimidines and an SBA and biochemical studies suggest that AB-423 likely also binds to the dimer-dimer interface of core protein. dual combination studies with AB-423 and anti-HBV agents, such as nucleos(t)ide analogs, RNA interference agents, or interferon alpha, resulted in additive to synergistic antiviral activity. Pharmacokinetic studies with AB-423 in CD-1 mice showed significant systemic exposures and higher levels of accumulation in the liver. A 7-day twice-daily administration of AB-423 in a hydrodynamic injection mouse model of HBV infection resulted in a dose-dependent reduction in serum HBV DNA levels, and combination with entecavir or ARB-1467 resulted in a trend toward antiviral activity greater than that of either agent alone, consistent with the results of the combination studies. The overall preclinical profile of AB-423 supports its further evaluation for safety, pharmacokinetics, and antiviral activity in patients with chronic hepatitis B.

Current approved nucleoside analogue treatments for chronic hepatitis B virus (HBV) infection are effective at controlling viral titer but are not curative and have minimal impact on the production of viral proteins such as surface antigen (HBsAg), the HBV envelope protein believed to play a role in maintaining the immune tolerant state required for viral persistence. Novel agents are needed to effect HBV cure, and reduction of HBV antigenemia may potentiate activation of effective and long-lasting host immune control. ARB-1740 is a clinical stage RNA interference agent composed of three siRNAs delivered using lipid nanoparticle technology. In a number of cell and animal models of HBV, ARB-1740 caused HBV RNA reduction, leading to inhibition of multiple elements of the viral life cycle including HBsAg, HBeAg, and HBcAg viral proteins as well as replication marker HBV DNA. ARB-1740 demonstrated pan-genotypic activity in vitro and in vivo, targeting three distinct highly conserved regions of the HBV genome, and effectively inhibited replication of nucleoside analogue-resistant HBV variants. Combination of ARB-1740 with a capsid inhibitor and pegylated interferon-alpha led to greater liver HBsAg reduction which correlated with more robust induction of innate immune responses in a human chimeric mouse model of HBV. The preclinical profile of ARB-1740 demonstrates the promise of RNA interference and HBV antigen reduction in treatment strategies driving toward a cure for HBV.

Telaprevir is a linear, peptidomimetic small molecule that inhibits hepatitis C virus (HCV) replication by specifically inhibiting the NS3·4A protease. In phase 3 clinical studies, telaprevir in combination with peginterferon and ribavirin (PR) significantly improved sustained virologic response (SVR) rates in genotype 1 chronic HCV-infected patients compared with PR alone. In patients who do not achieve SVR after treatment with telaprevir-based regimens, variants with mutations in the NS3·4A protease region have been observed. Such variants can contribute to drug resistance and limit the efficacy of treatment. To gain a better understanding of the viral resistance profile, we conducted phenotypic characterization of the variants using HCV replicons carrying site-directed mutations. The most frequently observed (significantly enriched) telaprevir-resistant variants, V36A/M, T54A/S, R155K/T, and A156S, conferred lower-level resistance (3-to 25-fold), whereas A156T and V36M؉R155K conferred higher-level resistance (>25-fold) to telaprevir. Rarely observed (not significantly enriched) variants included V36I/L and I132V, which did not confer resistance to telaprevir; V36C/G, R155G/I/M/S, V36A؉T54A, V36L؉R155K, T54S؉R155K, and R155T؉D168N, which conferred lower-level resistance to telaprevir; and A156F/N/V, V36A؉R155K/T, V36M؉R155T, V36A/ M؉A156T, T54A؉A156S, T54S؉A156S/T, and V36M؉T54S؉R155K, which conferred higher-level resistance to telaprevir. All telaprevir-resistant variants remained fully sensitive to alpha interferon, ribavirin, and HCV NS5B nucleoside and nonnucleoside polymerase inhibitors. In general, the replication capacity of telaprevir-resistant variants was lower than that of the wildtype replicon.

1 study. Variants with the substitutions L419C/I/M/P/S/V, R422K, M423I/T/V, I482L/N/T, A486S/T/V, and V494A were selected during VX-222 dosing, and their levels declined over time after the end of dosing. Phenotypic analysis of these variants was conducted using HCV replicons carrying site-directed mutations. Of the 17 variants, 14 showed reduced susceptibility to VX-222 compared with the wild type, with the L419C/S and R422K variants having higher levels of resistance (>200-fold) than the rest of the variants (6.8-to 76-fold). The M423I and A486S variants remained susceptible to VX-222. The 50% effective concentration (EC 50 ) for the L419P variant could not be obtained due to the poor replication of this replicon. The majority of the variants (15/17) were less fit than the wild type. A subset of the variants, predominately the L419S and R422K variants, were observed when the efficacy and safety of VX-222-and telaprevir-based regimens given for 12 weeks were investigated in genotype 1 HCV-infected patients in a phase 2 study. The NS3 and NS5B variants selected during the dual combination therapy showed reduced susceptibility to both telaprevir and VX-222 and had a lower replication capacity than the wild type. The phase 1b study has the ClinicalTrials.gov identifier NCT00911963, and the phase 2a study has ClinicalTrials.gov identifier NCT01080222.

We characterized a novel substitution conferring moderate resistance to telaprevir, a peptidomimetic inhibitor of hepatitis C virus protease. V36C conferred a 4.0-fold increase in the telaprevir 50% inhibitory concentration in an enzyme assay and a 9.5-fold increase in the replicon model. The replication capacity of a replicon harboring V36C was close to that of the wild-type protease. This case emphasizes the complexity of hepatitis C virus resistance to protease inhibitors.Advances in virology have led to the development of novel therapeutics specifically targeting hepatitis C virus (HCV) (4). Telaprevir (VX-950; Vertex Pharmaceuticals Incorporated, Cambridge, MA) is a novel, highly selective, potent peptidomimetic inhibitor of the HCV nonstructural protein 3/4A (NS3/4A) protease (1, 3) which has reached phase III clinical development in combination with pegylated alpha interferon (IFN-␣) and ribavirin. Amino acid substitutions conferring telaprevir resistance have been reported at positions Val 36, Thr 54, Arg 155, and Ala 156 of the NS3 protease (2, 5). In patients treated with telaprevir and pegylated IFN-␣ with and without ribavirin, breakthroughs during treatment and relapses after treatment are characterized by the recurrence of telaprevir-resistant HCV variant replication (1).Here, we characterized a novel, so far unknown, telaprevir resistance substitution at position Val 36 in a 38-year-old treatment-naïve woman with chronic hepatitis C due to HCV genotype 1b infection. The patient was enrolled in PROVE2, a phase II randomized clinical trial assessing the efficacy and safety of telaprevir in combination with pegylated IFN-␣2a with or without ribavirin (1). The patient was treated with telaprevir at 750 mg/8 h, pegylated IFN-␣2a at 180 g/week, and ribavirin at 1.0 g/day. HCV RNA became undetectable (Ͻ10 IU/ml) on therapy, but after 43 days of treatment, the patient withdrew consent and stopped therapy. She continued to be followed up after treatment withdrawal. Figure 1 shows the kinetics of HCV RNA levels in the patient during the 43 days of therapy. HCV RNA was detected 8 weeks after treatment withdrawal, and HCV RNA levels returned to nearly baseline levels. Twenty to 24 full-length NS3 protease clones were sequenced at each HCV RNA-positive time point. The patient was infected with a wild-type, telaprevir sensitive viral population at the baseline (Fig. 1). At the time of posttreatment relapse, the HCV variants all bore a Val-to-Cys substitution at position 36 (V36C). The V36C substitution remained dominant throughout posttreatment followup, up to day 512 after the start of therapy (Fig. 1). This substitution was associated with a Leu-to-Phe substitution at position 14 of the protease (L14F). L14F is present in approximately 6% of all HCV sequences and 8.5% of HCV subtype 1b sequences available in the European HCV database (http: //euhcvdb.ibcp.fr/euHCVdb/). Based on molecular modeling, it is predicted to be located at more than 20 Å of the telaprevir binding site and is therefore unlikely to alter...